U.S. Food and Drug Administration, Center for Veterinary Medicine, Office of Research, Division of Animal and Food Microbiology, 8401 Muirkirk Rd, Laurel, MD 20708, USA.
J AOAC Int. 2022 Oct 26;105(6):1503-1515. doi: 10.1093/jaoacint/qsac059.
Improvement in Salmonella detection methods greatly enhances the efficiency of various food testing programs. A Salmonella loop-mediated isothermal amplification (LAMP) assay has been validated in animal food through multi-laboratory validation.
The study aimed to demonstrate the versatility of this molecular assay while expanding it to multiple platforms and various reagent choices for use in animal food testing.
Following the U.S. Food and Drug Administration (FDA)'s Guidelines for the Validation of Analytical Methods for the Detection of Microbial Pathogens in Foods and Feeds, we examined the inclusivity, exclusivity, and LOD of the assay using two platforms (7500 Fast and Genie II) and three LAMP master mixes (GspSSD, GspSSD2.0, and WarmStart) in seven animal food matrixes (dry cat food, dry dog food, cattle feed, dairy feed, horse feed, poultry feed, and swine feed). The FDA's Bacteriological Analytical Manual (BAM) Salmonella culture method was the reference method.
Inclusivity and exclusivity data were consistent among all six platform and master mix combinations with a few exceptions. Comparable LODs were observed down to the single-cell level (WarmStart was 10-fold less sensitive). Performance was similar to the BAM method for detecting fractional positive results in seven animal food matrixes. Nonetheless, LAMP time to positive results and annealing/melting temperature differed among master mixes and platforms.
The Salmonella LAMP assay was successfully validated in two platforms and three master mixes, making it a flexible tool for use by the FDA's field laboratories in regulatory testing of animal food and for adoption by other food testing programs.
We demonstrated the LAMP assay's versatility on two platforms and three master mixes for the rapid and reliable screening of Salmonella in seven animal food matrixes. GspSSD2.0 was the fastest master mix (time to positive results as early as 3.5 min) while Genie II had several attractive features from a user perspective.
沙门氏菌检测方法的改进极大地提高了各种食品检测计划的效率。沙门氏菌环介导等温扩增（LAMP）检测方法已通过多实验室验证在动物食品中得到验证。
本研究旨在展示该分子检测方法的多功能性，同时将其扩展到多个平台和多种试剂选择，用于动物食品检测。
根据美国食品和药物管理局（FDA）的《食品和饲料中微生物病原体分析方法验证指南》，我们使用两种平台（7500 Fast 和 Genie II）和三种 LAMP 主混合物（GspSSD、GspSSD2.0 和 WarmStart）在七种动物食品基质（干粮、干粮、牛饲料、乳制品饲料、马饲料、家禽饲料和猪饲料）中检查了该检测方法的包容性、排他性和 LOD。FDA 的细菌分析手册（BAM）沙门氏菌培养方法是参考方法。
除了少数例外情况外，所有六种平台和主混合物组合的包容性和排他性数据均一致。在七种动物食品基质中，观察到可比的 LOD 低至单细胞水平（WarmStart 的灵敏度低 10 倍）。与 BAM 方法相比，该方法在七种动物食品基质中检测到分数阳性结果的性能相似。然而，LAMP 获得阳性结果的时间和退火/熔化温度因主混合物和平台而异。
沙门氏菌 LAMP 检测方法已在两种平台和三种主混合物中成功验证，使其成为 FDA 现场实验室在动物食品监管检测中以及其他食品检测计划中使用的灵活工具。
我们在两种平台和三种主混合物上展示了 LAMP 检测方法在七种动物食品基质中快速可靠筛选沙门氏菌的多功能性。GspSSD2.0 是最快的主混合物（最早 3.5 分钟即可获得阳性结果），而 Genie II 从用户角度具有几个吸引人的特点。
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